Proteomics/Genomics of Opiate Analgesia and Addiction

说明(申请人提供)：吗啡是一种高效的阿片类止痛药，是现代医学中一种有价值的止痛药。然而，长期使用吗啡的缺点是会发展成对耐受和依赖的上瘾。理想情况下，设计有效的止痛疗法将是有利的，因为它缺乏吗啡和相关阿片类药物的成瘾倾向。考虑到这一目标，有必要了解与耐受和依赖相比，介导吗啡镇痛的生化途径。当新的创新技术被应用于阐明具体的研究问题时，往往会出现科学进步。因此，将R21应用于先导性创新研究，利用蛋白质组学和基因组学的最新技术，将阐明与耐受或依赖相比，吗啡诱导镇痛所涉及的分子成分。值得注意的是，β-arrestin基因敲除小鼠在吗啡诱导的行为中表现出明显的变化，对应于增强的止痛，缺乏耐受性，而依赖性没有改变。这些发现导致了关于吗啡作用的这三个参数的不同生化途径的假说。因此，这个可行性项目的目标将是利用蛋白质组学和基因组学来比较慢性吗啡治疗期间β-arrestin基因敲除小鼠和野生型小鼠的调控分子成分，作为一种手段来阐明吗啡诱导镇痛与耐受或依赖相比的不同生化途径。具体目标是(1)与野生型小鼠相比，对慢性吗啡作用下大脑中受调控的蛋白质和多肽进行蛋白质组学评估；(2)使用独特的神经特异性小鼠基因芯片，在β-arrestin基因敲除小鼠和野生型小鼠中，对慢性吗啡过程中差异基因的表达进行基因组评估。这些研究将利用最先进的设施进行蛋白质组学和质谱学，并利用神经特异性小鼠基因芯片进行基因组学研究。将分析受调控的基因产物与基因家族的初级序列同源性，以预测其生物学功能。通过比较β-arrestin基因敲除小鼠和野生型小鼠的调控基因产物，可以将调控蛋白和基因分配给参与吗啡镇痛、耐受或依赖的候选生化途径。阐明不同的途径将具有重要意义，因为这可能使未来设计出提供有效止痛的阿片类药物方案，而不会对成瘾产生耐受和依赖。

DESCRIPTION (provided by applicant): Morphine is a highly effective opiate analgesic, and is a valuable painkiller in modern medicine. However, the disadvantage of chronic morphine administration is the development of addiction with tolerance and dependence. Ideally, it would be advantageous to design therapeutics for effective analgesia, which lacks the addictive liabilities of morphine and related opiates. With this goal in mind, it is necessary to understand the biochemical pathways that mediate morphine-induced analgesia compared to tolerance, and dependence. Scientific advances often occur when new innovative technologies are applied towards elucidating specific research issues. Therefore, this R21 application for pilot, innovative studies utilizing state-of-the-art technologies in both proteomics and genomics, will elucidate molecular components involved in morphine-induced analgesia compared to tolerance or dependence. Notably, beta-arrestin knockout mice show distinct changes in morphine-induced actions corresponding to enhanced analgesia, absence of tolerance, and no change in dependence. These findings lead to the hypothesis of distinct biochemical pathways for these three parameters of morphine actions. Therefore, the goal of this feasibility project will be to utilize both proteomics and genomics to compare regulated molecular components in beta-arrestin knockout mice and wild-type mice during chronic morphine treatment, as a means to elucidate distinct biochemical pathways for morphine-induced analgesia compared to tolerance or dependence. The specific aims are (1) proteomic evaluation of regulated proteins and peptides during chronic morphine in brains of beta-arrestin knockout mice compared to wild-type mice, and (2) genomic evaluation of differential gene expression during chronic morphine using unique neural-specific mouse cDNA microarrays in beta-arrestin knockout and wild-type mice. These studies will utilize state-of-the-art facilities for proteomics and mass spectrometry, and genomics with neural-specific mouse cDNA microarrays. Regulated gene products will be analyzed for primary sequence homology to gene families to predict their biological functions. Comparison of regulated gene products in beta-arrestin knockout and wild-type mice will allow assignment of regulated proteins and genes to candidate biochemical pathways involved in morphine analgesia, tolerance, or dependence. Elucidation of distinct pathways will be significant, since it could allow future design of opiate drug regimens that provide effective analgesia, without the tolerance and dependence of addiction.